Impact of low intensity pulsed ultrasound on volumetric root resorption of maxillary incisors in patients treated with clear aligner therapy: A retrospective study

ABSTRACT Objective: The aim of this study was to evaluate the volumetric root resorption in maxillary incisors following clear aligner therapy (CAT) with low-intensity pulsed ultrasound (LIPUS), and compare the results to CAT alone. Material and Methods: This retrospective study evaluated pretreatment (T0) and post-treatment (T1) cone-beam computed tomography imaging of 42 adult patients. Twenty-one patients (14 females, 7 males, mean age= 38.1±12.96 years) were treated using CAT with LIPUS device, whereas the other twenty-one matching controls patients (15 females, 6 males, mean age= 35.6±11.7 years) were treated using CAT alone. Images were analyzed and a segmentation protocol was applied on the maxillary incisors. Each segmented tooth volume was exported as a surface mesh in the Visualization Toolkit (VTK) file format. The VTK files for all maxillary incisors were coded and corresponding teeth volumes from T0 and T1 were superimposed. Clipping the crown of each tooth was done, then measurements of root volumes and differences between groups were performed. Changes in root volumes were assessed (p<0.05). Results: Root loss was evident in all teeth in both groups, but was significantly increased in all maxillary incisors of the control group (p<0.001) and in upper left central incisor of LIPUS group (p=0.009). When both groups were compared, there was statistically significant minimal volumetric root loss in LIPUS group (3.50-7.32 mm3), when compared to control group (11.48-12.95 mm3) (p<0.05). Conclusion: LIPUS group showed less volumetric root resorption compared to control group during the studied treatment time using clear aligners.

Impact of low intensity pulsed ultrasound on volumetric root resorption of maxillary incisors in patients treated with clear aligner therapy: A retrospective study INTRODUCTION Root resorption is an inevitable pathologic sequela of the biological processes that occur during orthodontic tooth movement (OTM).
This phenomenon is commonly known as orthodontically-induced inflammatory root resorption (OIIRR), and affects the surface of the tooth root -especially the apical part-, during which resorption of hard tissue components of the root may occurs. 1 OTM involves several biological processes and interactions at the cellular level along the root/bone interface from cementoenamel junction (CEJ) to root apex. Inflammatory reaction to the applied force is essential for OTM, but if uncontrolled, it triggers root resorption process. 2 The cellular mechanism of OIIRR is characterized by elevated concentrations of nuclear factor-kappa B ligand (RANKL) and reduced concentrations of osteoprotegerin (OPG). 3 It is well known that severe OIIRR compromises the success of orthodontic treatment. The maxillary incisors are the teeth most affected by OIIRR. 4 OIIRR is a complex multifactorial condition that is influenced by risk factors that may increase its incidence or severity: part of these factors is related to the patient, the others are related to the treatment mechanics. Patient-related risk factors includes abnormal root shape, 5-8 racial variation, 4 genetic predisposition, 9,10 being asthmatic 11 and hypodontia. 12 Several authors reported that there was no difference in either the incidence or severity of root resorption between male and female patients. 13 premolars to correct protruded teeth, 4,5,8 moving the apices for long distance, 6,7 lingual root torque, 16 intrusion of incisors, [17][18][19] intermaxillary elastics, 6 force magnitude 20 and long duration of the treatment. [16][17][18] Recent adjunctive interventions that aim to reduce total treatment duration and OIIRR are the main interest for several researchers. One type of these interventions is low-intensity pulsed ultrasound (LIPUS).
Ultrasound, an acoustic pressure wave at frequencies above the limit of human hearing, is transmitted through and into biologic tissues. It has been used widely in medicine as a therapeutic, operative, and diagnostic tool. 21 LIPUS output is of low intensity enough to be considered neither thermal nor destructive. 22 Previous research have shown that LIPUS reduces OIIRR through two major mechanisms: the first is stimulation of cementum deposition, through stimulation of cementoblasts; 23 and the second is inhibition of cementoclastogenesis, by altering OPG/RANKL ratio. 24 To date, no study assessed the effects of LIPUS on volumetric root loss in teeth after clear aligner therapy (CAT) in human subjects. The objective of this retrospective study was to assess if LIPUS reduces the severity of OIIRR in maxillary incisors, as evaluated by changes in tooth root volume after CAT, compared to no LIPUS treated patients. The null hypothesis tested was that there would be no significant difference in volumetric root loss in teeth treated using CAT with LIPUS or CAT alone.

MATERIAL AND METHODS
This was a retrospective study carried out on pretreatment (T0) and post-treatment (T1) cone beam computed tomography (CBCT) images of adult patients (age range between 18 years and 59 years) who were treated using CAT (Invisalign, Align Technology, Santa Clara, CA, USA) by the same orthodontist (TE) at his private orthodontic clinic in Edmonton, Canada, during a span of 5 years (2016-2020). CBCT images used were acquired as diagnostic records for orthodontic treatment planning. The patients had signed an informed consent form allowing the use of their data for scientific purposes. The study has been approved by the Health Research Ethics Board at the University of Alberta, Canada (Pro00091339). Data about sex, age, treatment duration and total number of aligners for each patient were also collected ( Table 1). During the orthodontic treatment planning stage, patients included in this study were instructed with information about tooth movement accelerating methods, in the form of brochures, videos and personalized discussions. The decision to use the LIPUS device or not was done by the patient and his/her family depending on their desire to shorten the treatment time and affordability for the extra cost of the adjunctive device. LIPUS was applied to the intervention group using an ultrasound device (Aevo system, SmileSonica Inc., Edmonton, AB, Canada) concurrently with CAT. The LIPUS device was used by the patient at home for 20 min/day during the whole treatment, with the parameters Al-Dboush R, Rossi A, El-Bialy T -Impact of low intensity pulsed ultrasound on volumetric root resorption of maxillary incisors in patients treated with clear aligner therapy: A retrospective study shown in Appendix 1. The other group, which served as a control group, was treated using CAT alone. The usage protocol of aligners in the intervention group was to change the aligners every 5 days, while the usage protocol of aligners in the control group was to change the aligners every 7 to 10 days.

LIPUS (n=21)
Control ( 26 After loading the DICOM data in the ITK-SNAP, a special tool called "Active Contour Segmentation Mode'' was used to select the area of interest (i.e., anterior maxilla) (Fig 1A). Following the selection of anterior part of the maxilla, 3-steps semi-automatic 3D segmentation wizard was started. The first step of the wizard is called pre-segmentation thresholding, being composed of two thresholds that helps the observer to visualize the image better by changing the values of gray scale: the maximum value of the upper thresholds was used, in order to include all the radiopaque structures; and the lower threshold was manipulated to obtain the most suitable gray value that showed good anatomy of the incisors. The next step was to place the baseline bubbles inside the maxillary incisors, which served as initiators for building up the 3D shape of the teeth ( Fig 1B). In the final step, which is called "evolution", actual contour segmentation was initiated, proceeded automatically in a stepwise fashion and manually stopped when the whole area of interest was covered by the colored labeling. Then, manual refinement of the teeth segmentation was undertaken using the Paintbrush mode on 2D image basis, to remove any voxels that represented surrounding anatomical structures (like bone plates or teeth other than maxillary incisors), and add any voxels that had been unintentionally omitted from the tooth volume during the semi-automatic segmentation process. Regarding the pulp cavity and canals, they were included during the manual refinement stage, to obtain the intact tooth and root volume. All refinements were performed on the multi-planar reformatted images, in axial, coronal, and sagittal orientation. After obttaining a 3D image of the four maxillary incisors as one labeled color, the scalpel tool was used in the 3D view screen, to assign a special color for each tooth. Once a special color was assigned for each tooth (Fig 1C), the "volumes and statistics" option was used to record the volume for each colored tooth structure in cubic

RESULTS
Characteristics of included patients in both groups are shown in

Overall, the mean root volumes for all incisors in both groups
were decreased at the end of orthodontic treatment ( Table 4). (Fig 3) showed that teeth of LIPUS group had statistically significant lower mean root volume loss, compared to the control group.

Results of comparison between the groups according to analysis of variance (independent samples Student's t-tests)
However, the difference was statistically significant in maxil-  respectively). The differences in treatment duration between the two groups were statistically significant (Table 1).

DISCUSSION
The aim of this retrospective study was to assess the possible effect of LIPUS on OIIRR during CAT using analysis of CBCT images at T0 and T1, and to assess the treatment efficiency of using LIPUS as an adjunctive to CAT.

Differences of root volumes [T1-T0 (mm 3 )] of the maxillary incisors in the LIPUS group
Upper right lateral incisor  (Table 3) showed large standard deviations, which may be due to patients' variability. Indeed, some teeth in the LIPUS group showed increase in the root volume at the end of treatment, as shown in Figure 4B. This was consistent with the results of previous studies 23,24,30 showing that LIPUS increases cementum formation, and this could have contributed to healing of root resorption with more cementum than the original shape of the teeth.
Regarding the total number of aligners used, there was no sta-  A limitation of this study was that the assessment of the outcomes was undertaken on a sample that did not include cases needing extractions. These cases are more susceptible to root resorption after moving the teeth to close the extraction spaces. The large field of view (FOV = 16x16 cm) and the voxel size of 0.3 mm were also considered limitations for this study. which leads to greater imaging noise, 36 and causes reduction in the spatial resolution 35 (i.e., the ability of the CBCT image to discriminate objects of different densities in close proximity). 37 Although it has been reported that the two most common voxel sizes used in orthodontics are 0.3 mm and 0.4 mm, 35 the results of one study showed that, for high precision volume measurements to be assessed accurately in-vivo, it would be better to choose a voxel size of 0.25 mm or less, as this makes the root segmentation process easier and increases the accuracy of the volume measurements obtained. However, better image quality requires a higher radiation dose and a longer scanning time. 38 Due to the retrospective nature of this study, other limitations would include the potential biases that could result from the patient's own decision on the group allocation due to additional treatment costs of using the LIPUS device, and the selection process of matched controls. Future randomized clinical trials can address these limitations. Finally, the lack of regression analysis test evaluating all variables that could affect root resorption and the lack of patient compliance reports of wearing the LIPUS device, that were not recorded, may be considered additional limitations for this study. Future studies with larger study sample, images with smaller FOV, and randomized design that includes extraction cases could validate the method that was used in this study by evaluating the volumetric OIIRR before extraction using the method mentioned above and compare it the volumetric OIIRR after extraction, by scanning the extracted teeth with a desktop micro-CT machine.

CONCLUSIONS
The null hypothesis of this study was rejected. Within the limits of this study, LIPUS daily use for 20 minutes could result in reduction of OIIRR extent when used in conjunction with CAT.
In general, this result may be related to the LIPUS capability of prevent or repair OIIRR and to the LIPUS-induced reduction of treatment time (which is a known risk factor for OIIRR).